This invention relates to a process for producing glucose/fructose syrups. More particularly, this invention relates to a process for producing glucose/fructose syrups comprising treating an unrefined glucose-containing starch hydrolysate with immobilized glucose isomerase.
Methods for producing glucose-containing starch hydrolysates are well known in the art and broadly fall into two categories: the acid-enzyme and enzyme-enzyme conversion processes. The latter process is generally preferred since it results in less reversion products being formed which are resistant to further treatment and therefore reduce the overall efficiency of the process.
In the enzyme-enzyme process an aqueous slurry is formed containing from about 30 to 40 percent dry substance starch and a starch digesting enzyme, typically bacterial alpha-amylase, is added thereto and the slurry heated to a temperature in the range of 80.degree. to 90.degree. C. to partially hydrolyze or liquefy the starch. Alpha-amylase is an endo-amylolytic enzyme capable of promoting random cleavage of .alpha.-1,4-glucosidic bonds within the starch molecule and is elaborated by a number of types of microorganisms, e.g., members of the Bacillus and Aspergillus genera, and also is present in malted cereal grains.
Alpha-amylase treatment results in only partial hydrolysis of the starch molecule since this enzyme does not act upon the .alpha.-1,6-glucosidic bonds in the molecule to a significant degree. Thus, alpha-amylase treated starch largely comprises oligosaccharides of varying molecular weights and fragments thereof which are more susceptible to further digestion by product-specific enzymes than is the untreated starch. To further hydrolyze the starch to provide a hydrolysate containing a significant proportion of glucose, the liquefied starch is treated with a glucogenic enzyme. Conventionally, the glucogenic enzyme employed is glucoamylase.
In the acid-enzyme process starch is first partially hydrolyzed or liquefied, e.g., by forming an aqueous suspension containing about 35 to 40 percent starch and incorporating therein an acid such as hydrochloric acid. The acidified suspension is then heated to high temperatures, cooled and treated with glucoamylase at a suitable concentration and pH to convert the partially hydrolyzed starch to glucose.
The use of glucose isomerase adsorbed onto or bonded to carriers to provide immobilized biological catalysts has largely superseded older methods whereby soluble enzymes or whole cells of microorganisms were utilized. In general, immobilized enzymes provide a number of advantages over these older methods, particularly in commercial systems for carrying out continuous conversion processes. Because of the economics involved in the utilization of such systems, it is of utmost importance that the stability or effective life of the immobilized enzyme be maintained over a period sufficient to permit conversion of large quantities of substrate. Methods of preparing immobilized glucose isomerase include bonding or otherwise adhering the enzyme to inert carriers such as derivatized cellulose, ion exchange resins and other polymeric materials, encapsulating the enzyme, entrapping the enzyme within fibers, etc.
Hitherto, attempts to use unrefined starch hydrolysates as substrates in continuous enzymatic processes for producing glucose/fructose syrups have not been efficient as desired due to the fact that the immobilized glucose isomerase becomes inactivated to a substantial degree after a relatively short period of use. This is believed to be due largely to the presence in the unrefined starch hydrolysate of materials which inhibit or otherwise deleteriously affect the activity of this enzyme.
Our investigation indicates that the stability or effective life of immobilized glucose isomerase is reduced by the presence in the unrefined starch hydrolysate of materials formed during the processes conventionally employed to liquefy and saccharify the starch. Although these materials have not been completely characterized, we believe that conditions for preparing starch hydrolysates which enhance the formation of such materials also tend to promote the non-enzymatic formation of ketose sugars, such as maltulose and fructose, or their precursors. The total concentration of either or both of these sugars, when produced by non-enzymatic action in an unrefined hydrolysate, therefore, can serve as an index of the suitability of the hydrolysate for enzymatic isomerization insofar as the prolongation of the activity of glucose isomerase is concerned.
It has hitherto been the general practice in the art to extensively refine or purify glucose-containing starch hydrolysates by known methods prior to isomerization of the glucose with glucose isomerase. Refining procedures commonly utilized include treatment of the clarified hydrolysate with carbon and ion exchange materials to remove undesirable constituents including metallic ions and carbohydrate degradation products.
Starch liquefaction processes are generally carried out at high temperatures in order to insure complete gelatinization of the starch granules. Liquefactions utilizing calcium dependent alpha-amylase preparations, e.g., those derived from B. subtilis, may require the presence of as much as 200 ppm of calcium ions, based on dry substance starch (dss), to impart optimum heat stability to this enzyme. Calcium in the form of lime has frequently been used for this purpose in starch liquefactions wherein it also serves to adjust the pH to the desired levels. However, the presence of substantial levels of calcium ions results in an undesirably high ash content in the hydrolysate and also, the calcium ion is a known inhibitor of glucose isomerase activity. Although it is probably impossible to avoid the presence of calcium altogether in starch hydrolysates prepared with alpha-amylase, for the purposes of the present invention, the hydrolysate should contain a concentration of not more than about 100 ppm of calcium ions, based on the content of the starch.
By carefully controlling the conditions under which the hydrolysate is prepared and avoiding those which promote the development therein of non-enzymatically generated ketose sugars as well as a high ash content, expensive procedures for refining or purifying the hydrolysate prior to isomerization with immobilized glucose isomerase may be eliminated with no significant reduction in the stability or effective life of the enzyme.